Testing of the antiviral effectiveness of the new and existing agents against Human Papillomavirus are still performed in the in vivo testing involving use of laboratory animals and human subjects. These studies are expensive, time consuming and altered by individual differences among subjects.
A demonstration of efficacy prior to in vivo animal model testing would limit the candidate in vivo agents to the ones with increasing potential for in vivo effectiveness. This is especially important for those newer, more speculative agents for which purer antiviral effects are lacking. In vitro demonstration of efficacy can support a decision for expensive testing in animal model systems. In vitro studies are also useful for exploring drug-virus interactions which are awkward or infeasible in whole animal systems. In vitro testing offers the following advantages: 1) preliminary data on efficacy; 2) rapid turn around time; 3) economy; 4) ability to precisely control environmental conditions; 5) elimination of pharmacokinetics and variability of whole animal systems; and 6) small amounts of drugs are required.
There are no established systems for in vitro papillomavirus testing. There have been some recent, encouraging developments, elsewhere and in our laboratory. Broker's laboratory has recently suggested that the xenograft system, which we originated, might be useful for antiviral testing (S. Dollard, et al., 1992, Gene Dev., 6:1131-1142).
In that one approach, fragments of HPV-11 infected human foreskin tissue is excised from the papillomatous cysts, growing beneath the renal capsule, and the fragments are placed onto a collagen gel "raft" culture. HPV-11 replication continues in the tissue fragment, as cells migrate laterally across the surface of the gel. We have explored the use of this system as a possible target for antiviral testing, and we have found that there is a high degree of regional variability in the extent of cell migration, tissue growth, and HPV-11 replication. We do not believe that this in vitro system is sufficiently consistent or precise to form a basis for tests. Further, since preliminary xenografts are required, the cost of the test includes their preliminary growth for three months, so some of the theoretical advantages of in vitro tests, economy and rapid turn-around are lacking.
Another in vitro system with potential was recently described by Laimins' group (Meyers, et al., 1992, Science, 257:971-973). In this system, human cervical cells, bearing HPV-31b episomal DNA are placed on collagen gel raft cultures and biosynthesis of complete virions occurs in the differentiating cells. It seems likely that this system may also be affected by regional variability.
Many of the disadvantages of the prior art methods of testing antiviral activity are overcome by the method of the present invention which precisely measures antiviral activity without the interferences of the regional variability, since the cell cultures are evenly dispersed monolayers.